X-ray structure and mechanism of RNA polymerase II stalled at an antineoplastic monofunctional platinum-DNA adduct

Abstract

DNA is a major target of anticancer drugs. The resulting adducts interfere with key cellular processes, such as transcription, to trigger downstream events responsible for drug activity. cisDiammine(pyridine)chloroplatinum(II), cDPCP or pyriplatin, is a monofunctional platinum(II) analogue of the widely used anticancer drug cisplatin having significant anticancer properties with a different spectrum of activity. Its novel structure-activity properties hold promise for overcoming drug resistance and improving the spectrum of treatable cancers over those responsive to cisplatin. However, the detailed molecular mechanism by which cells process DNA modified by pyriplatin and related monofunctional complexes is not at all understood. Here we report the structure of a transcribing RNA polymerase II (pol II) complex stalled at a site-specific monofunctional pyriplatin-DNA adduct in the active site. The results reveal a molecular mechanism of pol II transcription inhibition and drug action that is dramatically different from transcription inhibition by cisplatin and UV-induced 1,2-intrastrand cross-links. Our findings provide insight into structure-activity relationships that may apply to the entire family of monofunctional DNA-damaging agents and pave the way for rational improvement of monofunctional platinum anticancer drugs.anticancer | chemotherapy | DNA damage | pyriplatin | transcription T he DNA template for transcription is not only the site of inborn errors of metabolism and of continuous attack by harmful environmental agents, but it also represents a major target for cancer therapy. Platinum-based anticancer drugs such as cisplatin, cis-diamminedichloroplatinum(II), are widely used and among the most effective antineoplastic treatments (1, 2). Platinum-based drugs typically form bifunctional intra-or interstrand DNA cross-links by covalent bonding to the N 7 positions of two guanosine residues, triggering a variety of cellular processes, including transcription inhibition with attendant apoptosis (1, 2). However, resistance and side effects can require withdrawal of these drugs before they can effect a cure in certain types of cancer (3).In the effort to find new compounds that circumvent resistance to conventional bifunctional platinum-based drugs, a class of monofunctional platinum compounds were synthesized and screened for anticancer activity (4-6). In contrast to other inactive monofunctional platinum(II) compounds such as ½PtðdienÞCl þ and ½PtðNH 3 Þ 3 Cl þ , cis-diammine(pyridine)chloroplatinum(II) [cDPCP or "pyriplatin" (Fig. 1)] and related complexes display significant anticancer properties and a different spectrum of activity compared to conventional platinum-based drugs. These features render them attractive candidates for treating cisplatin-refractory patients if the potency could be raised to or beyond the level of that of cisplatin (4, 5, 7). Pyriplatin exhibits unique chemical and biological properties, forming monofunctional DNA adducts ( Fig. 1 and Fig. S1) that can inhibit transcription an...

Millisecond dynamics of RNA polymerase II translocation at atomic resolution

“…Finally (state 3 to state 4), the transition base rotates to the canonical position of posttranslocation state, which allows full base pairing with the incoming NTP. The intermediate state's identity is consistent with previous translocation intermediate structures trapped either by α-amanitin or DNA damage (13,37), in which the TN base is located on top of the BH.…”

“…To proceed to the next NAC, Pol II must translocate from a pretranslocation state, in which the active site is still occupied by the newly added nucleotide at 3′-RNA, to a posttranslocation state. During translocation, the template DNA and RNA must move by exactly one register, once again creating a free insertion site (i site) (1,2,5,(8)(9)(10)(11)(12)(13).…”

Third row transition metals for the treatment of cancer

“…A structural analysis of RNA pol II stalled at a site-specific pyriplatin lesion suggested that, by increasing the steric bulk of the N-heterocyclic ligand, it might be possible to increase transcription inhibition and consequential cytotoxicity [32]. From a panel of different N-heterocyclic ligands, Am, a series of complexes having the formula cis-[Pt(NH 3 ) 2 Cl(Am)] + was prepared [33].…”

RNA polymerase II acts as a selective sensor for DNA lesions and endogenous DNA modifications

“…Aligning the structures of 5caC-paused pol II elongation complex with several recent bulky DNA lesionarrested complexes reveals new insights into pol II pausing and arrest upon DNA modifications/ lesions. 12,17,26,29 Intriguingly, the iC1 transition template base in these 5caC-paused or bulky DNA lesion-arrested structures are all accommodated "above the bridge helix," representing a common translocation checkpoint for a variety of endogenous DNA modifications and DNA lesions. 12,17,26,29 Intriguingly, the similar "above the bridge helix" translocation intermediates are also observed for non-damaged pausing and arrest complexes revealed by the structures of the a-amanitinarrested pol II complex 5,30 and the E. Coli RNAP pausing complex.…”

“…12,17,26,29 Intriguingly, the iC1 transition template base in these 5caC-paused or bulky DNA lesion-arrested structures are all accommodated "above the bridge helix," representing a common translocation checkpoint for a variety of endogenous DNA modifications and DNA lesions. 12,17,26,29 Intriguingly, the similar "above the bridge helix" translocation intermediates are also observed for non-damaged pausing and arrest complexes revealed by the structures of the a-amanitinarrested pol II complex 5,30 and the E. Coli RNAP pausing complex. 31 Another interesting example is revealed by a recent structure of the RNA pol III elongation complex, 32 in which the undamaged template iC1 nucleobase adopts a similar "above the bridge helix" position.…”